Actin is the most abundant protein in animal cells and constitutes 10-20% of the protein of many nucleated cells and 30% of the protein of muscle cells. Actin molecules each bind an ATP molecule and self-assemble into long filaments during which the ATP is hydrolyzed into ADP.
Injury to animal tissues results in the release of actin into the extracellular space, including the bloodstream. Although approximately half of nonmuscle cell actin is F-actin, (the double-helical, rodlike, filament form of actin which is assembled from G-actin monomers), the ionic conditions of extracellular fluids favor actin polymerization, so that virtually all the actin released into the blood from dying cells would be expected to polymerize into filaments (Lind, S. E. et al., Am. Rev. Respir. Dis. 138:429-434 (1988)). In purified solutions, in the absence of filament-shortening proteins, actin filaments can easily attain lengths of several microns. Were some fraction of actin released from injured cells to be irreversibly denatured, however, or else bound to one of the intracellular actin-binding proteins discussed below, this actin would remain monomeric.
There are many proteins which naturally associate with actin (for a review of actin-binding proteins see Stossel, T. P. et al., Ann. Rev. Cell Biol. 1:353-402 (1985); Pollard, T. D. et al., Ann. Rev. Biochem. 55:987-1035 (1986)). However, two proteins, gelsolin and DBP (vitamin D binding protein) are thought to be primarily responsible for binding extracellular actin (Janmey, P. A. et al., Blood 70:529-530 (1987)).
Gelsolin has been cloned (Kwiatkowski, D. J. et al., Nature 323:455-458 (1986); Kwiatkowski, D. J. et al., J. Cell Biol. 106:375-384 (1988)) and fragments of the native protein which retain the ability to bind actin have been identified (Bryan, J., J. Cell Biol. 106:1553-1562 (1988); Yin, H. L. et al,., J. Cell Biol. 107:465a (1988), abst. no. 2616); Kwiatkowski, D. J. et al., J. Cell Biol. 108:1717-1726 (1989); Way, M. et al., J. Cell Biol. 109:593-605 (1989)). DBP has also been cloned (Cooke, N. E. et al., J. Clin. Invest. 76:2420-2424 (1985); Yang, F. et al., Proc. Natl. Acad. Sci. U.S.A. 82:7994-7998 (1985)).
Because of the large amounts of actin in cells, the release of actin from dying cells provides sufficient actin to have a significant affect on the microenvironment, either by increasing the viscosity of extracellular fluids of plasma and/or by entrapping cells or by other, as yet unidentified toxic effects. Infusion of extracellular free actin is toxic to animal tissues, and especially to renal and cardiopulmonary systems (Harper, K. D. et al., Clin. Res. 36:625A (1988); Haddad, J. G. et al., Proc. Natl. Acad. Sci. U.S.A. 87:1381-1385 (1990)). Acute renal failure is a complication of muscle injury and actin infusions in rats causes transient elevations of the BUN and creatinine levels, consistent with renal failure.
Moreover, since each extracellular actin molecule in a filament has an ADP molecule associated with it, the presence of extracellular actin in the blood may tend to induce or augment platelet aggregation in a manner which may not be advantageous to the host (Lind, S. E. et al., Am. Rev. Respir. Dis 138:429-434 (1988); Scarborough et al., Biochem. Biophy. Res. Commun. 100:1314-1319 (1981)).
It has been proposed that the binding of extracellular actin to gelsolin or DBP protects against F-actin formation in the circulation and may be the mechanism by which such actin is targeted for removal from the bloodstream (Lind, S. E. et. al., J. Clin. Invest. 78:736-742 (1986); Sanger, J. et al., Clin. Res. 36:625A (1988)). For example, it is known that circulating actin-gelsolin complexes are found following oleic-acid induced lung injury in rabbits and cats (Smith, D. B. et al., Am. J. Path. 130:261-267 (1988)) and in phenylhydrazine-induced hemolysis in rabbits (Smith, D. B. et al., Blood 72:214-218, 1988)). In the clinical setting, gelsolin levels are depressed in patients with the adult respiratory distress syndrome (ARDS) (Lind, S. E. et al., Am. Rev. Respir. Dis. 138:429-434 (1988)), and in the plasma of patients with acute falciparum malaria infection (Smith, D. B. et al., Blood 72:214-218 (1988)), and gelsolin actin complexes are detectable in the blood of such patients. In addition, complexes of actin with DBP have been observed in the plasma of patients with fulminant hepatic necrosis (Young, W. O. et al., J. Lab. Clin. Med. 110:83 (1987)). It has also been proposed that once the ability of plasma proteins to bind extracellular free actin is exceeded, intravascular filament formation and endothelial injury can be detected (Harper, K. D. et al., Clin. Res. 36:625A (1988); Haddad, J. G. et al., Proc. Natl. Acad. Sci. 87:1381-1389 (1990)).
It is known that actin impairs the normal sequence of blood coagulation and fibrinolytic events. Actin inhibits the normal transition of fibrin from a "fine" to a "course" clot (Janmey, P. A. et al., Biochim. Biophys. Acta 841:151-161 (1985)), and is also a potent inhibitor of the fibrinolytic enzyme plasmin.
A treatment which can prevent or ameliorate tissue injury in a subject which occurs in response to actin in the bloodstream and/or tissues has not been described. Hence, a need exists for a method which can ameliorate, treat or prevent such tissue injury.